Our objective is to identify and optimize plant derived small molecules for the inhibition of HIV RT-associated RNase H (RNH) avtivity, a novel target for HIV intervention. The overall goal of this poject is to recommend 2-3 compounds for development (CD) from the initial screen of 10,000 fractions (1st program) and recommend 4-6 CDs (back up program) from screening the rest of the plant derived library (150,000 fractions). The first generation CDs should attain an IC50 of 50 nm or better while second generation CDs should have single digit nm IC50. These compounds should be effective in ihibiting HIV strains that are resistant to current drugs. RNH inhibitors (RNHI) will be optimized using biochemical and virologic data initially followed by utilizing computanional and structural information obtained from X-ray crystallographic studies of RT-RNHI complexes (see other projects within this RFA) to allow rational design of analogs that will be either synthesized or semi-synthesized. In case a semi-synthetic approach is adopted, we can use our 50 L bioreactor to produce gram quantities of the starting material to allow us to generate a small library of analogs. In addition, combinatorial and parallel synthesis will be utilized for some molecules to increase both the quantity and chemical diversity of potential leads. We will adhere to established guidlines such as the Lipinski "Rule of five" to increase our chances of obtaining drug like molecules. Analogs that show improved acivity will be subjected to ADME/PK studies in order to select for drug like leads. This process will be repeated until we achieve the profile of a CD described in Table 4. We would like to include yet another approach in our drug discovery of RNHI. We will exploit the generation of metabolites of some of our leads as a source of new chemotypes for identifyiing RNHI. We are very encouraged by the results that we obtained so far (Tablel). There is a strong correlation between the biochemical and cell-based values, some having an IC50<1uM with an excellent therapeutic Index >200. This represents a major step forward in identifying and developing RNHIs. It is a tribute to the plant cell culture technology that we developed so far. A highly diverse library that originated from more than 2200 plant species, representing 284 plant families, that is charecarized by a high reproducibility of secondary metabolite production. This project has enormous potential impact in delivering a new class of HIV drugs.